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Comparative studies on the possible origin of extremely high contents of vitamin D_3 in some kinds of fish liver were undertaken. When bastared halibuts and carps were farmed from fingerlings to adults with feedstuffs containing vitamin D_2 or D_3, significant amounts of the vitamins were accumulated in the fish liver. Significant amounts of vitamin D_2 and D_3 in phytoplankton and vitamin D_3 in zooplankton were detected. Therefore, we concluded that the most probable origin of vitamin D_3 in fish liver is a result of food chains from plankton. Vitamin D metabolism in fish was investigated. When [^3H]-25-OH-D_3 was incubated with liver homogenates of D-deficient carp, a peak corresponding to [^3H]-1,25 (OH)_2D_3 was observed in the profiles of HPLC. Although 25-OH-D_3-1α-hydroxylase was also observed in the kidney, the activity of the enzyme was lower than that in the liver. Therefore, we concluded that fish contain 25-OH-D_3-1α-hydroxylase in the liver besides in the kidney. Comparative studies on vitamin D metabolism in vertebrates were undertaken. The formation of 1,25 (OH)_2D_3 was detected in the liver of fish and amphibians but not reptiles and mammals, while 1,25(OH)_2D_3 was formed in all of the kidney. The existence of vitamin D-binding protein (DBP) specifically bound to 25-OH-D_3 was not found in the plasma of fish and amphibians while it was in those of reptiles and mammals. These results suggest that in development of a special transporting system of 25-OH-D_3 using plasma DBP, the main 25-OH-D_3 -1α-hydroxylase activity changes from the liver to the kidney. We could confirm the existence of 25-OH-D_3-1α-hydroxylase in the liver of fetal rats in addition to that in the kidney. This indicates "ontogeny is a recapitulation of phylogeny" with respect to vitamin D metabolism.

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Aromatic _L-amino acid decarboxylase (AADC) decarboxylates both _L-5-hydroxytryptophan to serotonin in serotonergic neurons and pineal cells, and _L-DOPA to dopamine in catecholaminergic neurons and adrenal medullary cells. Thus AADC produces two major mammalian neurotransmitters and hormones. We isolated and sequenced a full-length, neuronal-type, cDNA encoding human AADC. It consisted of 1932 bases containing an open reading frame encoding 480 amino acids residues with a molecular weight of 53,891. We expressed a recobinant human AADC in COS cells and proved that the expressed enzyme decarboxylated both _L-5-hydroxytryptophan to serotonin and _L-DOPA to dopamine. We cloned genomic DNA of human AADC and determined the structure. The genomic DNA of human AADC consists of 15 exons spanning about 100 kilobases and exists as a single copy in the haploid genome. We have mapped the gene to chromosome band 7p12.1-p12.3 by fluorescene in situ hybridization. We cloned the nonneuronal type cDNA from human liver and identified another first exon different from the neuronal type cDNA. This showed that an alternative usage of the first exon produced two types of mRNAs in AADC and suggested that alternative splicing would regulate the tissue-specific expression of AADC.

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In the in vitro lipid peroxidation dependent on microsomal cytochrome P-450, the production of thiobarbituric acid-reactive substance (TBARS) was increase by the addition of ethanol in the reaction mixture. The increase of TBARS was inhibited by the addition of flavin adenine dinucleotide (FAD), superoxide dismutase or catalase, and was also inhibited by uric acid or mannitol but not inhibited by sodium azide. The reaction of p-nitroso-N, N-dimethylaniline and hydroxyl radical (・OH) was inhibited by FAD, and FAD was decreased by the reaction of ・OH production. These results suggest that the active oxygen radical, such as ・OH, generates by the metabolism of ethanol in microsome reacted more rapidly with FAD than unsaturated fatty acid, and then the lipid peroxidation is inhibited.

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We examined effects of 1α-OH-D_3 on bone loss in ovariectomized rats. Since bone mineral density of spines and femurs were decreased by ovariectomy, 1α-OH-D_3 significantly prevented bone loss in a dose dependent manner. 1α-OH-D_3 also significantly kept bone strength in femur to the normal levels. These effects were similar to those of 1,25(OH)_2D_3. Bone morphometric study showed stimulated bone resorption in the rats decreased by the administrations of 1α-OH-D_3. These results were suggestive that 1α-OH-D_3 shows preventions effects on osteoporosis due to inhibiting stimulated bone turn-over in the patients.

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